In recent years, much has been learned about megakaryocytopoiesis and the roles played by thrombopoietin and its receptor, Mp1. Certain diseases have been associated with the failure of TPO signaling (i.e. thrombocytopenia with absent radii (TAR) and amegakaryocytic thrombocytopenia) as well as excessive thrombopoietin production (i.e. familial thrombocytosis). In addition, an important role for thrombopoietin in stem cell and primitive progenitor production has been recognized. The potent proliferative stimulus delivered by TPO may play a role in the etiology or progression of myeloid leukemia as those cancers that express Mp1 have a worse prognosis than those that are negative for Mp1 expression. The specific aims of this proposal build upon our recent work towards a goal of understanding the molecular signaling mechanisms that are essential for megakaryocyte proliferation and differentiation. A more complete understanding of these processes may improve our ability to expand hematopoietic stem cells or produce megakaryocytes and/or platelets in vitro. Furthermore, the ability to disrupt specific proliferative signaling pathways may lead to novel therapeutic approaches to myeloid leukemias. The specific aims for this proposal include the following: 1) Study the structure/function relationships and signaling pathways involved in cellular proliferation that arise from the membrane-proximal region of Mp1 . 2) Determine the internalization signal that facilitates ligand-dependent internalization of Mp1. 3) Examine how Src kinases are utilized during megakaryocyte development. 4) Determine the basic biological differences between megakaryocytes derived from umbilical cord blood and adult bone marrow. These studies will help answer important questions about hematopoiesis and the role of cytokines in regulating proliferation and lineage-specific differentiation.